Veterans are particularly susceptible to neuropathic pain originating from combat wounds, surgery and diabetes. The long-term goal of this project is to find a cure for neuropathic pain that represents its true reversal and not just palliative measures. This goal can be achieved by investigating the molecular and cellular mechanisms of pain latent sensitization, a rodent model that reproduces key features of chronic pain disorders: indefinite duration and episodic presentation triggered by stress. Latent sensitization can occur following a variety of stimuli: inflammation, skin incision and nerve injuries that cause neuropathic pain. It is characterized by a period of hyperalgesia followed by a period of pain remission in which pain is generally absent but may reappear as pain episodes. This is due to the fact that the remission phase of latent sensitization does not represent a return to normality but an anomalous state in which pain is suppressed by a compensatory activation of opioid receptors. This is revealed by a return of pain (?reinstatement?) upon administration of opioid antagonists, which are without effect in normal animals or humans. The latent sensitization model emerged from the work of several research teams on opiate-induced hyperalgesia and its interaction with injury and stress. Only recently has the importance of latent sensitization as a model for chronic pain become clear, and there is only one study on its relevance for neuropathic pain. This project will investigate the mechanisms of latent sensitization in neuropathic pain. The Specific Aims are: 1) study the role of opioid receptors and stress in neuropathic latent sensitization; 2) determine whether the remission phase of latent sensitization is mediated by opioid release or by -opioid receptor constitutive activity; and 3) study the role of NMDARs and the adenylyl cyclase / protein kinase A pathway in neuropathic latent sensitization. We will test the following hypotheses: 1) -opioid receptors, ?-opioid receptors and ?2A adrenergic receptors in the spinal cord mediate anti-hyperalgesia during the remission phase of nerve injury-induced latent sensitization; 2) -opioid receptors located in primary afferent terminals contribute to the remission phase of latent sensitization; 3) stress reinstates pain hyperalgesia during the remission phase of latent sensitization; 4) -opioid receptor constitutive activity, and not sustained opioid release, mediates pain remission; 5) pain sensitivity in latent sensitization is maintained by a positive feedback loop in which NMDARs and the adenylyl cyclase / protein kinase A pathway activate each other. This project will use a multidisciplinary approach to test these hypotheses. In Aim 1, a spared nerve injury model of neuropathic pain will be used in rats and mice to identify the receptors that suppress pain sensitivity in the remission phase. Using transgenic mice with selective deletion of -opioid receptors in nociceptive primary afferents, we will determine whether the -opioid receptors that suppress pain are located in primary afferents. We will also investigate how stress reinstates pain during the remission phase. In Aim 2, we will determine whether the anti-hyperalgesia produced by -opioid receptors is due to their constitutive activity by using patch-clamp recording from primary afferent neurons to measure their inhibition of voltage-gated calcium channels. The alternative hypothesis, that -opioid receptors are activated by sustained opioid release, will be tested using knock-out mice in the three genes that encode opioid peptides. In addition, opioid release will be measured as -opioid receptor internalization. In Aim 3, the interaction between NMDA receptors, adenylyl cyclase and protein kinase A during latent sensitization will be studied in primary afferent neurons using calcium imaging and patch-clamp. The activating phosphorylation of NMDA receptors by protein kinase A during latent sensitization will be measured with Western blots. Finally, NMDA receptor antagonists and inhibitors and activators of adenylyl cyclase and protein kinase A will be used to delineate the signal pathway that maintains latent sensitization. This will identify targets to develop medication to reverse neuropathic pain.